Genetic and Phenotypic Characterization of a Rabies Virus Strain Isolated from a Dog in Tokyo, Japan in the 1940s.

The rabies virus strain Komatsugawa (Koma), which was isolated from a dog in Tokyo in the 1940s before eradication of rabies in Japan in 1957, is known as the only existent Japanese field strain (street strain). Although this strain potentially provides a useful model to study rabies pathogenesis, little is known about its genetic and phenotypic properties. Notably, this strain underwent serial passages in rodents after isolation, indicating the possibility that it may have lost biological characteristics as a street strain. In this study, to evaluate the utility of the Koma strain for studying rabies pathogenesis, we examined the genetic properties and in vitro and in vivo phenotypes. Genome-wide genetic analyses showed that, consistent with previous findings from partial sequence analyses, the Koma strain is closely related to a Russian street strain within the Arctic-related phylogenetic clade. Phenotypic examinations in vitro revealed that the Koma strain and the representative street strains are less neurotropic than the laboratory strains. Examination by using a mouse model demonstrated that the Koma strain and the street strains are more neuroinvasive than the laboratory strains. These findings indicate that the Koma strain retains phenotypes similar to those of street strains, and is therefore useful for studying rabies pathogenesis.

Development of infectious clones of a wild-type Korean rabies virus and evaluation of their pathogenic potential.

Most reverse genetic (RG) systems for rabies viruses (RVs) have been constructed on the genome background of laboratory-adapted strains. In this study, we developed an RG system using a Korean wild type (KGH) strain to investigate the pathogenic potential of different strains. We developed a RG system with the KGH strain for the first time. Following the complete genome sequencing of the KGH strain, pKGH infectious clones were constructed using the CMV/T7 promoter, and HamRz and HdvRz were introduced to allow self-cleavage of the synthesized RNA. We successfully recovered the rescued virus by constructing chimeric RVs in which we replaced a part of the construct with the partial gene from the fixed RC-HL strain. The rescued viruses formed clearer and countable plaques in an immunostaining plaque assay, with a distinct plaque morphology. Furthermore, compared with the chimeric RVs, the pKGH/RCinsΔ4 strain containing the KGH strain G protein exhibited a decreased efficiency of cell-to-cell spreading in BHK-21 cells and significantly reduced (100-1000 fold) replication kinetics. However, pKGH/RCinsΔ4 strain-infected mice revealed 100% morbidity at 11days post-infection, whereas other chimeric RV strains showed no mortality. Our RG system is a useful tool for studying differences in the cell-to-cell spreading efficiency and replication with respect to the different internalization patterns of street and fixed laboratory-adapted viruses.